bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2022‒01‒02
twenty-two papers selected by
Kelsey Fisher-Wellman
East Carolina University
  1. Computational modeling of mitochondrial K+- and H+-driven ATP synthesis.
  2. Defining the interactome of the human mitochondrial ribosome identifies SMIM4 and TMEM223 as respiratory chain assembly factors.
  3. Interfering with mitochondrial dynamics sensitizes glioblastoma multiforme to temozolomide chemotherapy.
  4. Induction and Detection of Mitophagy.
  5. Mitochondrial superoxide targets energy metabolism to modulate epigenetic regulation of NRF2-mediated transcription.
  6. SOD2 V16A Amplifies Vascular Dysfunction in Sickle Cell Patients by Curtailing Mitochondria Complex IV Activity.
  7. Mitochondria in human neutrophils mediate killing of Staphylococcus aureus.
  8. Patient-derived iPSCs link elevated mitochondrial respiratory complex I function to osteosarcoma in Rothmund-Thomson syndrome.
  9. Ultrastructural changes in cristae of lymphoblasts in acute lymphoblastic leukemia parallel alterations in biogenesis markers.
  10. Mitochondrial Ndufa4l2 Enhances Deposition of Lipids and Expression of Ca9 in the TRACK Model of Early Clear Cell Renal Cell Carcinoma.
  11. The mitochondrial DNA constitution shaping T-cell immunity in patients with rectal cancer at high risk of metastatic progression.
  12. SIRT2 regulates mitochondrial dynamics and reprogramming via MEK1-ERK-DRP1 and AKT1-DRP1 axes.
  13. Model systems in SDHx-related pheochromocytoma/paraganglioma.
  14. AMPK-mTOR-mediated activation of autophagy promotes formation of dormant polyploid giant cancer cells.
  15. Macrophages are metabolically heterogeneous within the tumor microenvironment.
  16. NAD kinase sustains lipogenesis and mitochondrial metabolismthrough fatty acid synthesis.
  17. Sod1 integrates oxygen availability to redox regulate NADPH production and the thiol redoxome.
  18. pH-dependent 11° F1FO ATP synthase sub-steps reveal insight into the FO torque generating mechanism.
  19. NDUFC1 Is Upregulated in Gastric Cancer and Regulates Cell Proliferation, Apoptosis, Cycle and Migration.
  20. Overexpression of carnitine palmitoyltransferase 1A promotes mitochondrial fusion and differentiation of glioblastoma stem cells.
  21. Redox-Sensitive VDAC: A Possible Function as an Environmental Stress Sensor Revealed by Bioinformatic Analysis.
  22. A cell culture platform for quantifying metabolic substrate oxidation in bicarbonate-buffered medium.
  1. J Mol Cell Cardiol. 2021 Dec 23. pii: S0022-2828(21)00232-7. [Epub ahead of print]165 9-18
    Computational modeling of mitochondrial K+- and H+-driven ATP synthesis.
    Sonia Cortassa, Miguel A Aon, Magdalena Juhaszova, Evgeny Kobrinsky, Dmitry B Zorov, Steven J Sollott.
      ATP synthase (F1Fo) is a rotary molecular engine that harnesses energy from electrochemical-gradients across the inner mitochondrial membrane for ATP synthesis. Despite the accepted tenet that F1Fo transports exclusively H+, our laboratory has demonstrated that, in addition to H+, F1Fo ATP synthase transports a significant fraction of ΔΨm-driven charge as K+ to synthesize ATP. Herein, we utilize a computational modeling approach as a proof of principle of the feasibility of the core mechanism underlying the enhanced ATP synthesis, and to explore its bioenergetic consequences. A minimal model comprising the 'core' mechanism constituted by ATP synthase, driven by both proton (PMF) and potassium motive force (KMF), respiratory chain, adenine nucleotide translocator, Pi carrier, and K+/H+ exchanger (KHEmito) was able to simulate enhanced ATP synthesis and respiratory fluxes determined experimentally with isolated heart mitochondria. This capacity of F1Fo ATP synthase confers mitochondria with a significant energetic advantage compared to K+ transport through a channel not linked to oxidative phosphorylation (OxPhos). The K+-cycling mechanism requires a KHEmito that exchanges matrix K+ for intermembrane space H+, leaving PMF as the overall driving energy of OxPhos, in full agreement with the standard chemiosmotic mechanism. Experimental data of state 4➔3 energetic transitions, mimicking low to high energy demand, could be reproduced by an integrated computational model of mitochondrial function that incorporates the 'core' mechanism. Model simulations display similar behavior compared to the experimentally observed changes in ΔΨm, mitochondrial K+ uptake, matrix volume, respiration, and ATP synthesis during the energetic transitions at physiological pH and K+ concentration. The model also explores the role played by KHEmito in modulating the energetic performance of mitochondria. The results obtained support the available experimental evidence on ATP synthesis driven by K+ and H+ transport through the F1Fo ATP synthase.
    Keywords:  Energy supply-demand matching; F(1)F(o) ATP synthase; Mitochondrial K(+) uptake; Mitochondrial K(+)/H(+) exchanger
    DOI:  https://doi.org/10.1016/j.yjmcc.2021.12.005
  2. Elife. 2021 Dec 31. pii: e68213. [Epub ahead of print]10
    Defining the interactome of the human mitochondrial ribosome identifies SMIM4 and TMEM223 as respiratory chain assembly factors.
    Sven Dennerlein, Sabine Poerschke, Silke Oeljeklaus, Cong Wang, Ricarda Richter-Dennerlein, Johannes Sattmann, Diana Bauermeister, Elisa Hanitsch, Stefan Stoldt, Thomas Langer, Stefan Jakobs, Bettina Warscheid, Peter Rehling.
      Human mitochondria express a genome that encodes thirteen core subunits of the oxidative phosphorylation system (OXPHOS). These proteins insert into the inner membrane co-translationally. Therefore, mitochondrial ribosomes engage with the OXA1L-insertase and membrane-associated proteins, which support membrane insertion of translation products and early assembly steps into OXPHOS complexes. To identify ribosome-associated biogenesis factors for the OXPHOS system, we purified ribosomes and associated proteins from mitochondria. We identified TMEM223 as a ribosome-associated protein involved in complex IV biogenesis. TMEM223 stimulates the translation of COX1 mRNA and is a constituent of early COX1 assembly intermediates. Moreover, we show that SMIM4 together with C12ORF73 interacts with newly synthesized cytochrome b to support initial steps of complex III biogenesis in complex with UQCC1 and UQCC2. Our analyses define the interactome of the human mitochondrial ribosome and reveal novel assembly factors for complex III and IV biogenesis that link early assembly stages to the translation machinery.
    Keywords:  assembly; biochemistry; cell biology; chemical biology; mitochondria; oxidative phosphorylation; ribosome; translation
    DOI:  https://doi.org/10.7554/eLife.68213
  3. J Cell Mol Med. 2021 Dec 28.
    Interfering with mitochondrial dynamics sensitizes glioblastoma multiforme to temozolomide chemotherapy.
    Nan Wang, Renxuan Huang, Kunmeng Yang, Yichun He, Yufei Gao, Delu Dong.
      Glioblastoma multiforme (GBM) is a primary tumour of the central nervous system (CNS) that exhibits the highest degree of malignancy. Radiotherapy and chemotherapy are essential to prolong the survival time of patients. However, clinical work has demonstrated that sensitivity of GBM to chemotherapy decreases with time. The phenomenon of multi-drug resistance (MDR) reminds us that there may exist some fundamental mechanisms in the process of chemo-resistance. We tried to explore the mechanism of GBM chemo-resistance from the perspective of energy metabolism. First, we found that the oxidative phosphorylation (OXPHOS) level of SHG44 and U87 cells increased under TMZ treatment. In further studies, it was found that the expression of PINK1 and mitophagy flux downstream was downregulated in GBM cells, which were secondary to the upregulation of TP53 in tumour cells under TMZ treatment. At the same time, we examined the mitochondrial morphology in tumour cells and found that the size of mitochondria in tumour cells increased under the treatment of TMZ, which originated from the regulation of AMPK on the subcellular localization of Drp1 under the condition of unbalanced energy supply and demand in tumour cells. The accumulation of mitochondrial mass and the optimization of mitochondrial quality accounted for the increased oxidative phosphorylation, and interruption of the mitochondrial fusion process downregulated the efficiency of oxidative phosphorylation and sensitized GBM cells to TMZ, which was also confirmed in the in vivo experiment. What is more, interfering with this process is an innovative strategy to overcome the chemo-resistance of GBM cells.
    Keywords:  AMPK; TP53; glioblastoma multiforme; mitochondrial dynamics; temozolomide
    DOI:  https://doi.org/10.1111/jcmm.17147
  4. Methods Mol Biol. 2022 ;2445 227-239
    Induction and Detection of Mitophagy.
    Maria A Yapryntseva, Boris Zhivotovsky, Vladimir Gogvadze.
      Mitophagy, a process of selective elimination of mitochondria by autophagy, is a mechanism of mitochondrial quality control that maintains mitochondrial network functionality. The elimination of damaged mitochondria through autophagy requires two steps: induction of general autophagy and priming of damaged mitochondria for selective autophagic recognition. Mitophagy impairment is linked to various pathologies; thus, removal of malfunctioning or even harmful mitochondria is vital to cellular physiology. Here, we describe methods that can be applied to the investigation of mitophagy.
    Keywords:  Autophagy; Confocal microscopy; Flow cytometry; Mitochondria; Mitophagy; Respiration
    DOI:  https://doi.org/10.1007/978-1-0716-2071-7_14
  5. Free Radic Biol Med. 2021 Dec 27. pii: S0891-5849(21)01169-2. [Epub ahead of print]
    Mitochondrial superoxide targets energy metabolism to modulate epigenetic regulation of NRF2-mediated transcription.
    Sanjit K Dhar, Timothy Scott, Chi Wang, Teresa W M Fan, Daret K St Clair.
      Mitochondria are central to the metabolic circuitry that generates superoxide radicals/anions (O2•-) as a by-product of oxygen metabolism. By regulating superoxide levels, manganese superoxide dismutase plays important roles in numerous biochemical and molecular events essential for the survival of aerobic life. In this study, we used MitoParaquat (mPQ) to generate mitochondria-specific O2•- and stable isotope-resolved metabolomics tracing in primary human epidermal keratinocytes to investigate how O2•- generated in mitochondria regulates gene expression. The results reveal that isocitrate is blocked from conversion to α-ketoglutarate and that acetyl-coenzyme A (CoA) accumulates, which is consistent with a reduction in oxygen consumption rate and inactivation of isocitrate dehydrogenase (IDH) activity. Since acetyl-CoA is linked to histone acetylation and gene regulation, we determined the effect of mPQ on histone acetylation. The results demonstrate an increase in histone H3 acetylation at lysines 9 and 14. Suppression of IDH increased histone acetylation, providing a direct link between metabolism and epigenetic alterations. The activity of histone acetyltransferase p300 increased after mPQ treatment, which is consistent with histone acetylation. Importantly, mPQ selectively increased the nuclear levels and activity of the oxidative stress-sensitive nuclear factor erythroid 2-related factor 2. Together, the results establish a new paradigm that recognizes O2•- as an initiator of metabolic reprogramming that activates epigenetic regulation of gene transcription in response to mitochondrial dysfunction.
    Keywords:  Epigenetics; Metabolism; Mitochondria; Superoxide; TCA cycle; Transcription
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2021.12.309
  6. Blood. 2021 Dec 27. pii: blood.2021013350. [Epub ahead of print]
    SOD2 V16A Amplifies Vascular Dysfunction in Sickle Cell Patients by Curtailing Mitochondria Complex IV Activity.
    Atinuke Dosunmu-Ogunbi, Shuai Yuan, Michael Reynolds, Luca Giordano, Subramaniam Sanker, Mara Sullivan, Donna B Stolz, Brett A Kaufman, Katherine C Wood, Yingzhe Zhang, Sruti Shiva, Seyed Mehdi Nouraie, Adam C Straub.
      Superoxide dismutase 2 (SOD2) catalyzes the dismutation of superoxide to hydrogen peroxide in mitochondria limiting mitochondrial damage. The SOD2 amino acid valine-to-alanine substitution at position 16 (V16A) in the mitochondrial leader sequence is a common genetic variant among patients with sickle cell disease (SCD). However, little is known about the cardiovascular consequences of SOD2V16A in SCD patients or its impact on endothelial cell function. Here, we show SOD2V16A associates with increased tricuspid regurgitant velocity (TRV), systolic blood pressure, right ventricle area at systole and declined 6-minute walk distance in 410 SCD patients. Plasma lactate dehydrogenase, a marker of oxidative stress and hemolysis, significantly associated with higher TRV. To define the impact of SOD2V16A in the endothelium, we introduced the SOD2V16A variant into endothelial cells. SOD2V16A increases hydrogen peroxide and mitochondrial reactive oxygen species (ROS) production compared to controls. Unexpectedly, the increased ROS was not due to SOD2V16A mislocalization but was associated with mitochondrial Complex IV and a concomitant decrease in basal respiration and Complex IV activity. In sum, SOD2V16A is a novel clinical biomarker of cardiovascular dysfunction in SCD patients through its ability to decrease mitochondrial Complex IV activity and amplify ROS production in the endothelium.
    DOI:  https://doi.org/10.1182/blood.2021013350
  7. Redox Biol. 2021 Dec 24. pii: S2213-2317(21)00385-2. [Epub ahead of print]49 102225
    Mitochondria in human neutrophils mediate killing of Staphylococcus aureus.
    Kimberly J Dunham-Snary, Bas Gj Surewaard, Jeffrey D Mewburn, Rachel Et Bentley, Ashley Y Martin, Oliver Jones, Ruaa Al-Qazazi, Patricia Ad Lima, Paul Kubes, Stephen L Archer.
      BACKGROUND: Neutrophils play a role in innate immunity and are critical for clearance of Staphylococcus aureus. Current understanding of neutrophil bactericidal effects is that NADPH oxidase produces reactive oxygen species (ROS), mediating bacterial killing. Neutrophils also contain numerous mitochondria; since these organelles lack oxidative metabolism, their function is unclear. We hypothesize that mitochondria in human neutrophils contribute to the bactericidal capacity of S. aureus.METHODS: and Findings: Using human neutrophils isolated from healthy volunteers (n = 13; 7 females, 6 males), we show that mitochondria are critical in the immune response to S. aureus. Using live-cell and fixed confocal, and transmission electron microscopy, we show mitochondrial tagging of bacteria prior to ingestion and surrounding of phagocytosed bacteria immediately upon engulfment. Further, we demonstrate that mitochondria are ejected from intact neutrophils and engage bacteria during vital NETosis. Inhibition of the mitochondrial electron transport chain at Complex III, but not Complex I, attenuates S. aureus killing by 50 ± 7%, comparable to the NADPH oxidase inhibitor apocynin. Similarly, mitochondrial ROS scavenging using MitoTEMPO attenuates bacterial killing 112 ± 60% versus vehicle control. Antimycin A treatment also reduces mitochondrial ROS production by 50 ± 12% and NETosis by 53 ± 5%.
    CONCLUSIONS: We identify a previously unrecognized role for mitochondria in human neutrophils in the killing of S. aureus. Inhibition of electron transport chain Complex III significantly impairs antimicrobial activity. This is the first demonstration that vital NETosis, an early event in the antimicrobial response, occurring within 5 min of bacterial exposure, depends on the function of mitochondrial Complex III. Mitochondria join NADPH oxidase as bactericidal ROS generators that mediate the bactericidal activities of human neutrophils.
    Keywords:  Electron transport chain complex III; Immunity; Neutrophil extracellular trap (NET); Nicotinamide adenine dinucleotide phosphate (NADPH) oxidases; Phagocytosis; Staphylococcus aureus
    DOI:  https://doi.org/10.1016/j.redox.2021.102225
  8. PLoS Genet. 2021 Dec;17(12): e1009971
    Patient-derived iPSCs link elevated mitochondrial respiratory complex I function to osteosarcoma in Rothmund-Thomson syndrome.
    Brittany E Jewell, An Xu, Dandan Zhu, Mo-Fan Huang, Linchao Lu, Mo Liu, Erica L Underwood, Jun Hyoung Park, Huihui Fan, Julian A Gingold, Ruoji Zhou, Jian Tu, Zijun Huo, Ying Liu, Weidong Jin, Yi-Hung Chen, Yitian Xu, Shu-Hsia Chen, Nino Rainusso, Nathaniel K Berg, Danielle A Bazer, Christopher Vellano, Philip Jones, Holger K Eltzschig, Zhongming Zhao, Benny Abraham Kaipparettu, Ruiying Zhao, Lisa L Wang, Dung-Fang Lee.
      Rothmund-Thomson syndrome (RTS) is an autosomal recessive genetic disorder characterized by poikiloderma, small stature, skeletal anomalies, sparse brows/lashes, cataracts, and predisposition to cancer. Type 2 RTS patients with biallelic RECQL4 pathogenic variants have multiple skeletal anomalies and a significantly increased incidence of osteosarcoma. Here, we generated RTS patient-derived induced pluripotent stem cells (iPSCs) to dissect the pathological signaling leading to RTS patient-associated osteosarcoma. RTS iPSC-derived osteoblasts showed defective osteogenic differentiation and gain of in vitro tumorigenic ability. Transcriptome analysis of RTS osteoblasts validated decreased bone morphogenesis while revealing aberrantly upregulated mitochondrial respiratory complex I gene expression. RTS osteoblast metabolic assays demonstrated elevated mitochondrial respiratory complex I function, increased oxidative phosphorylation (OXPHOS), and increased ATP production. Inhibition of mitochondrial respiratory complex I activity by IACS-010759 selectively suppressed cellular respiration and cell proliferation of RTS osteoblasts. Furthermore, systems analysis of IACS-010759-induced changes in RTS osteoblasts revealed that chemical inhibition of mitochondrial respiratory complex I impaired cell proliferation, induced senescence, and decreased MAPK signaling and cell cycle associated genes, but increased H19 and ribosomal protein genes. In summary, our study suggests that mitochondrial respiratory complex I is a potential therapeutic target for RTS-associated osteosarcoma and provides future insights for clinical treatment strategies.
    DOI:  https://doi.org/10.1371/journal.pgen.1009971
  9. Appl Microsc. 2021 Dec 29. 51(1): 20
    Ultrastructural changes in cristae of lymphoblasts in acute lymphoblastic leukemia parallel alterations in biogenesis markers.
    Ritika Singh, Ayushi Jain, Jayanth Kumar Palanichamy, T C Nag, Sameer Bakhshi, Archna Singh.
      We explored the link between mitochondrial biogenesis and mitochondrial morphology using transmission electron microscopy (TEM) in lymphoblasts of pediatric acute lymphoblastic leukemia (ALL) patients and compared these characteristics between tumors and control samples. Gene expression of mitochondrial biogenesis markers was analysed in 23 ALL patients and 18 controls and TEM for morphology analysis was done in 15 ALL patients and 9 healthy controls. The area occupied by mitochondria per cell and the cristae cross-sectional area was observed to be significantly higher in patients than in controls (p-value = 0.0468 and p-value< 0.0001, respectively). The mtDNA copy numbers, TFAM, POLG, and c-myc gene expression were significantly higher in ALL patients than controls (all p-values< 0.01). Gene Expression of PGC-1α was higher in tumor samples. The analysis of the correlation between PGC-1α expression and morphology parameters i.e., both M/C ratio and cristae cross-sectional area revealed a positive trend (r = 0.3, p = 0.1). The increased area occupied by mitochondria and increased cristae area support the occurrence of cristae remodelling in ALL. These changes might reflect alterations in cristae dynamics to support the metabolic state of the cells by forming a more condensed network. Ultrastructural imaging can be useful for affirming changes occurring at a subcellular organellar level.
    Keywords:  Acute lymphoblastic leukemia; Cristae; Electron microscopy; Mitochondrial biogenesis; Mitochondrial morphology
    DOI:  https://doi.org/10.1186/s42649-021-00069-4
  10. Front Oncol. 2021 ;11 783856
    Mitochondrial Ndufa4l2 Enhances Deposition of Lipids and Expression of Ca9 in the TRACK Model of Early Clear Cell Renal Cell Carcinoma.
    Kristian B Laursen, Qiuying Chen, Francesca Khani, Nabeel Attarwala, Steve S Gross, Lukas Dow, David M Nanus, Lorraine J Gudas.
      Mitochondrial dysfunction and aberrant glycolysis are hallmarks of human clear cell renal cell carcinoma (ccRCC). Whereas glycolysis is thoroughly studied, little is known about the mitochondrial contribution to the pathology of ccRCC. Mitochondrial Ndufa4l2 is predictive of poor survival of ccRCC patients, and in kidney cancer cell lines the protein supports proliferation and colony formation. Its role in ccRCC, however, remains enigmatic. We utilized our established ccRCC model, termed Transgenic Cancer of the Kidney (TRACK), to generate a novel genetically engineered mouse model in which dox-regulated expression of an shRNA decreases Ndufa4l2 levels specifically in the renal proximal tubules (PT). This targeted knockdown of Ndufa4l2 reduced the accumulation of neutral renal lipid and was associated with decreased levels of the ccRCC markers carbonic anhydrase 9 (CA9) and Enolase 1 (ENO1). These findings suggest a link between mitochondrial dysregulation (i.e. high levels of Ndufa4l2), lipid accumulation, and the expression of ccRCC markers ENO1 and CA9, and demonstrate that lipid accumulation and ccRCC development can potentially be attenuated by inhibiting Ndufa4l2.
    Keywords:  HIF1; clear cell renal cell carcinoma; hypoxia; lipids; mitochondria; proximal tubules; von Hippel-Lindau
    DOI:  https://doi.org/10.3389/fonc.2021.783856
  11. Clin Transl Oncol. 2021 Dec 27.
    The mitochondrial DNA constitution shaping T-cell immunity in patients with rectal cancer at high risk of metastatic progression.
    P A Bousquet, S Meltzer, A J Fuglestad, T Lüders, Y Esbensen, H V Juul, C Johansen, L G Lyckander, T Bjørnetrø, E M Inderberg, C Kersten, K R Redalen, A H Ree.
      PURPOSE: A significant percentage of colorectal cancer patients proceeds to metastatic disease. We hypothesised that mitochondrial DNA (mtDNA) polymorphisms, generated by the high mtDNA mutation rate of energy-demanding clonal immune cell expansions and assessable in peripheral blood, reflect how efficiently systemic immunity impedes metastasis.PATIENTS AND METHODS: We studied 44 rectal cancer patients from a population-based prospective biomarker study, given curative-intent neoadjuvant radiation and radical surgery for high-risk tumour stage and followed for metastatic failure. Blood specimens were sampled at the time of diagnosis and analysed for the full-length mtDNA sequence, composition of immune cell subpopulations and damaged serum mtDNA.
    RESULTS: Whole blood total mtDNA variant number above the median value for the study cohort, coexisting with an mtDNA non-H haplogroup, was representative for the mtDNA of circulating immune cells and associated with low risk of a metastatic event. Abundant mtDNA variants correlated with proliferating helper T cells and cytotoxic effector T cells in the circulation. Patients without metastatic progression had high relative levels of circulating tumour-targeting effector T cells and, of note, the naïve (LAG-3+) helper T-cell population, with the proportion of LAG-3+ cells inversely correlating with cell-free damaged mtDNA in serum known to cause antagonising inflammation.
    CONCLUSION: Numerous mtDNA polymorphisms in peripheral blood reflected clonal expansion of circulating helper and cytotoxic T-cell populations in patients without metastatic failure. The statistical associations suggested that patient's constitutional mtDNA manifests the helper T-cell capacity to mount immunity that controls metastatic susceptibility.
    TRIAL REGISTRATION: ClinicalTrials.gov NCT01816607; registration date: 22 March 2013.
    Keywords:  CD4; Colorectal cancer; Immune cells; Metastasis; Mitochondrial DNA
    DOI:  https://doi.org/10.1007/s12094-021-02756-w
  12. Cell Rep. 2021 Dec 28. pii: S2211-1247(21)01651-X. [Epub ahead of print]37(13): 110155
    SIRT2 regulates mitochondrial dynamics and reprogramming via MEK1-ERK-DRP1 and AKT1-DRP1 axes.
    Young Cha, Taewoo Kim, Jeha Jeon, Yongwoo Jang, Patrick B Kim, Claudia Lopes, Pierre Leblanc, Bruce M Cohen, Kwang-Soo Kim.
      During somatic reprogramming, cellular energy metabolism fundamentally switches from predominantly mitochondrial oxidative phosphorylation toward glycolysis. This metabolic reprogramming, also called the Warburg effect, is critical for the induction of pluripotency, but its molecular mechanisms remain poorly defined. Notably, SIRT2 is consistently downregulated during the reprogramming process and regulates glycolytic switch. Here, we report that downregulation of SIRT2 increases acetylation of mitogen-activated protein kinase (MAPK) kinase-1 (MEK1) at Lys175, resulting in activation of extracellular signal-regulated kinases (ERKs) and subsequent activation of the pro-fission factor dynamin-related protein 1 (DRP1). In parallel, downregulation of SIRT2 hyperacetylates the serine/threonine protein kinase AKT1 at Lys20 in a non-canonical way, activating DRP1 and metabolic reprogramming. Together, our study identified two axes, SIRT2-MEK1-ERK-DRP1 and SIRT2-AKT1-DRP1, that critically link mitochondrial dynamics and oxidative phosphorylation to the somatic reprogramming process. These upstream signals, together with SIRT2's role in glycolytic switching, may underlie the Warburg effect observed in human somatic cell reprogramming.
    Keywords:  AKT1; DRP1; MEK1-ERK axis; OXPHOS; SIRT2; Warburg-like effect; human somatic cell reprogramming; induced pluripotent stem cells; metabolic reprogramming; mitochondrial remodeling
    DOI:  https://doi.org/10.1016/j.celrep.2021.110155
  13. Cancer Metastasis Rev. 2021 Dec 27.
    Model systems in SDHx-related pheochromocytoma/paraganglioma.
    Krisztina Takács-Vellai, Zsolt Farkas, Fanni Ősz, Gordon W Stewart.
      Pheochromocytoma (PHEO) and paraganglioma (PGL) (together PPGL) are tumors with poor outcomes that arise from neuroendocrine cells in the adrenal gland, and sympathetic and parasympathetic ganglia outside the adrenal gland, respectively. Many follow germline mutations in genes coding for subunits of succinate dehydrogenase (SDH), a tetrameric enzyme in the tricarboxylic acid (TCA) cycle that both converts succinate to fumarate and participates in electron transport. Germline SDH subunit B (SDHB) mutations have a high metastatic potential. Herein, we review the spectrum of model organisms that have contributed hugely to our understanding of SDH dysfunction. In Saccharomyces cerevisiae (yeast), succinate accumulation inhibits alpha-ketoglutarate-dependent dioxygenase enzymes leading to DNA demethylation. In the worm Caenorhabditis elegans, mutated SDH creates developmental abnormalities, metabolic rewiring, an energy deficit and oxygen hypersensitivity (the latter is also found in Drosophila melanogaster). In the zebrafish Danio rerio, sdhb mutants display a shorter lifespan with defective energy metabolism. Recently, SDHB-deficient pheochromocytoma has been cultivated in xenografts and has generated cell lines, which can be traced back to a heterozygous SDHB-deficient rat. We propose that a combination of such models can be efficiently and effectively used in both pathophysiological studies and drug-screening projects in order to find novel strategies in PPGL treatment.
    Keywords:  Model organisms; Paraganglioma; Pheochromocytoma; SDH; Succinate dehydrogenase
    DOI:  https://doi.org/10.1007/s10555-021-10009-z
  14. Cancer Res. 2021 Dec 29. pii: canres.2342.2021. [Epub ahead of print]
    AMPK-mTOR-mediated activation of autophagy promotes formation of dormant polyploid giant cancer cells.
    Bo You, Tian Xia, Miao Gu, Zhenxin Zhang, Qicheng Zhang, Jianhong Shen, Yue Fan, Hui Yao, Si Pan, Yingna Lu, Tianyi Cheng, Zhiyuan Yang, Xin He, Hao Zhang, Muqi Shi, Dong Liu, Yiwen You.
      Dormant cancer cells that survive anti-cancer therapy can lead to cancer recurrence and disseminated metastases that prove fatal in most cases. Recently, specific dormant polyploid giant cancer cells (PGCC) have drawn our attention because of their association with the clinical risk of nasopharyngeal carcinoma (NPC) recurrence, as demonstrated by previous clinical data. In this study, we report the biological properties of PGCC, including mitochondrial alterations, and reveal that autophagy is a critical mechanism of PGCC induction. Moreover, pharmacological or genetic inhibition of autophagy greatly impaired PGCC formation, significantly suppressing metastasis and improving survival in a mouse model. Mechanistically, chemotherapeutic drugs partly damaged mitochondria, which then produced low ATP levels and activated autophagy via the AMPK-mTOR pathway to promote PGCC formation. Analysis of the transcriptional and epigenetic landscape of PGCC revealed overexpression of RIPK1, and the scaffolding function of RIPK1 was required for AMPK-mTOR pathway-induced PGCC survival. High numbers of PGCCs correlated with shorter recurrence time and worse survival outcomes in NPC patients. Collectively, these findings suggest a therapeutic approach of targeting dormant PGCCs in cancer.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-21-2342
  15. Cell Rep. 2021 Dec 28. pii: S2211-1247(21)01671-5. [Epub ahead of print]37(13): 110171
    Macrophages are metabolically heterogeneous within the tumor microenvironment.
    Xenia Geeraerts, Juan Fernández-Garcia, Felix J Hartmann, Kyra E de Goede, Liesbet Martens, Yvon Elkrim, Ayla Debraekeleer, Benoit Stijlemans, Anke Vandekeere, Gianmarco Rinaldi, Riet De Rycke, Mélanie Planque, Dorien Broekaert, Elisa Meinster, Emile Clappaert, Pauline Bardet, Aleksandar Murgaski, Conny Gysemans, Frank Aboubakar Nana, Yvan Saeys, Sean C Bendall, Damya Laoui, Jan Van den Bossche, Sarah-Maria Fendt, Jo A Van Ginderachter.
      Macrophages are often prominently present in the tumor microenvironment, where distinct macrophage populations can differentially affect tumor progression. Although metabolism influences macrophage function, studies on the metabolic characteristics of ex vivo tumor-associated macrophage (TAM) subsets are rather limited. Using transcriptomic and metabolic analyses, we now reveal that pro-inflammatory major histocompatibility complex (MHC)-IIhi TAMs display a hampered tricarboxylic acid (TCA) cycle, while reparative MHC-IIlo TAMs show higher oxidative and glycolytic metabolism. Although both TAM subsets rapidly exchange lactate in high-lactate conditions, only MHC-IIlo TAMs use lactate as an additional carbon source. Accordingly, lactate supports the oxidative metabolism in MHC-IIlo TAMs, while it decreases the metabolic activity of MHC-IIhi TAMs. Lactate subtly affects the transcriptome of MHC-IIlo TAMs, increases L-arginine metabolism, and enhances the T cell suppressive capacity of these TAMs. Overall, our data uncover the metabolic intricacies of distinct TAM subsets and identify lactate as a carbon source and metabolic and functional regulator of TAMs.
    Keywords:  TCA cycle break; immunometabolism; immunosuppression; lactate; macrophage metabolism; metabolomics; non-small-cell lung carcinoma; single-cell metabolic profiling; tumor microenvironment; tumor-associated macrophages
    DOI:  https://doi.org/10.1016/j.celrep.2021.110171
  16. Cell Rep. 2021 Dec 28. pii: S2211-1247(21)01653-3. [Epub ahead of print]37(13): 110157
    NAD kinase sustains lipogenesis and mitochondrial metabolismthrough fatty acid synthesis.
    Mengyao Xu, Long Ding, Jingjing Liang, Xiao Yang, Yuan Liu, Yingchun Wang, Mei Ding, Xun Huang.
      Lipid storage in fat tissue is important for energy homeostasis and cellular functions. Through RNAi screening in Drosophila fat body, we found that knockdown of a Drosophila NAD kinase (NADK), which phosphorylates NAD to synthesize NADP de novo, causes lipid storage defects. NADK sustains lipogenesis by maintaining the pool of NADPH. Promoting NADPH production rescues the lipid storage defect in the fat body of NADK RNAi animals. Furthermore, NADK and fatty acid synthase 1 (FASN1) regulate mitochondrial mass and function by altering the levels of acetyl-CoA and fatty acids. Reducing the level of acetyl-CoA or increasing the synthesis of cardiolipin (CL), a mitochondrion-specific phospholipid, partially rescues the mitochondrial defects of NADK RNAi. Therefore, NADK- and FASN1-mediated fatty acid synthesis coordinates lipid storage and mitochondrial function.
    Keywords:  Drosophila; FASN; NADK; lipogenesis; mitochondrial metabolism
    DOI:  https://doi.org/10.1016/j.celrep.2021.110157
  17. Proc Natl Acad Sci U S A. 2022 Jan 04. pii: e2023328119. [Epub ahead of print]119(1):
    Sod1 integrates oxygen availability to redox regulate NADPH production and the thiol redoxome.
    Claudia Montllor-Albalate, Hyojung Kim, Anna E Thompson, Alex P Jonke, Matthew P Torres, Amit R Reddi.
      Cu/Zn superoxide dismutase (Sod1) is a highly conserved and abundant antioxidant enzyme that detoxifies superoxide (O2 •-) by catalyzing its conversion to dioxygen (O2) and hydrogen peroxide (H2O2). Using Saccharomyces cerevisiae and mammalian cells, we discovered that a major aspect of the antioxidant function of Sod1 is to integrate O2 availability to promote NADPH production. The mechanism involves Sod1-derived H2O2 oxidatively inactivating the glycolytic enzyme, GAPDH, which in turn reroutes carbohydrate flux to the oxidative phase of the pentose phosphate pathway (oxPPP) to generate NADPH. The aerobic oxidation of GAPDH is dependent on and rate-limited by Sod1. Thus, Sod1 senses O2 via O2 •- to balance glycolytic and oxPPP flux, through control of GAPDH activity, for adaptation to life in air. Importantly, this mechanism for Sod1 antioxidant activity requires the bulk of cellular Sod1, unlike for its role in protection against O2 •- toxicity, which only requires <1% of total Sod1. Using mass spectrometry, we identified proteome-wide targets of Sod1-dependent redox signaling, including numerous metabolic enzymes. Altogether, Sod1-derived H2O2 is important for antioxidant defense and a master regulator of metabolism and the thiol redoxome.
    Keywords:  glycolysis; oxygen sensing; pentose phosphate pathway; redox signaling; superoxide dismutase
    DOI:  https://doi.org/10.1073/pnas.2023328119
  18. Elife. 2021 Dec 31. pii: e70016. [Epub ahead of print]10
    pH-dependent 11° F1FO ATP synthase sub-steps reveal insight into the FO torque generating mechanism.
    Seiga Yanagisawa, Wayne D Frasch.
      Most cellular ATP is made by rotary F1FO ATP synthases using proton translocation-generated clockwise torque on the FO c-ring rotor, while F1-ATP hydrolysis can force counterclockwise rotation and proton pumping. The FO torque-generating mechanism remains elusive even though the FO interface of stator subunit-a, which contains the transmembrane proton half-channels, and the c-ring is known from recent F1FO structures. Here, single-molecule F1FO rotation studies determined that the pKa values of the half-channels differ, show that mutations of residues in these channels change the pKa values of both half-channels, and reveal the ability of FO to undergo single c-subunit rotational stepping. These experiments provide evidence to support the hypothesis that proton translocation through FO operates via a Grotthuss mechanism involving a column of single water molecules in each half-channel linked by proton translocation-dependent c-ring rotation. We also observed pH-dependent 11° ATP synthase-direction sub-steps of the E. coli c10-ring of F1FO against the torque of F1-ATPase-dependent rotation that result from H+ transfer events from FO subunit-a groups with a low pKa to one c-subunit in the c-ring, and from an adjacent c-subunit to stator groups with a high pKa. These results support a mechanism in which alternating proton translocation-dependent 11° and 25° synthase-direction rotational sub-steps of the c10-ring occur to sustain F1FO ATP synthesis.
    Keywords:  E. coli; molecular biophysics; structural biology
    DOI:  https://doi.org/10.7554/eLife.70016
  19. Front Oncol. 2021 ;11 709044
    NDUFC1 Is Upregulated in Gastric Cancer and Regulates Cell Proliferation, Apoptosis, Cycle and Migration.
    Liang Xu, Xiuxiu Chen, Hongtao Jiang, Jian Xu, Lixia Wang, Yuemin Sun.
      Gastric cancer is one of the most common primary tumors of the digestive system. NADH: ubiquinone oxidoreductase subunit C1 (NDUFC1), which is an accessory subunit of the NADH dehydrogenase (complex I), is responsible for the transportation of electrons from NADH to the respiratory chain essential for the oxidative phosphorylation. However, little is known about the roles of NDUFC1 in carcinogenesis. In this study, NDUFC1 protein level in NSCLC tissues was tested by immunohistochemistry (IHC) staining. NDUFC1 mRNA level in gastric cancer cell lines was determined by qRT-PCR. MGC-803 and SGC-7901 cells were transfected with shNDUFC1 lentivirus designed to silence NDUFC1. MTT assay, CCK8 assay, wound healing assay and transwell migration assay were conducted. Cell cycle and apoptosis were detected by flow cytometry. In vivo experiments were performed using nude mice. The results indicated that overexpressed NDUFC1 in gastric cancer was related to more serious tumor infiltrates, a higher risk of lymphatic metastasis, a higher proportion of positive lymph nodes, and a more advanced tumor stage. Compared with shCtrl groups, MGC-803 and SGC-7901 of shNDUFC1 groups had lower abilities of proliferation and migration, higher levels of apoptosis. NDUFC1 knockdown also inhibited SGC-7901 cell growth in vivo and suppressed Ki67 expression in xenograft tumors. More importantly, we found that NDUFC1 downregulation made the levels of P-Akt, P-mTOR, CCND1, CDK6, PIK3CA, Bcl-2, Survivin, and XIAP decreased, and that PI3K/AKT signaling pathway agonist SC79 rescued the inhibitory effects on cell proliferation and migration, reversed the promoted effects on cell apoptosis caused by NDUFC1 knockdown. More importantly, compared with NDUFC1 knockdown group, the expression of P-Akt, Bcl-2, Survivin, and XIAP was raised in shNDUFC1 + SC79 group. Thus, our suspicion was that NDUFC1 exacerbates NSCLC progression via PI3K/Akt pathway. Taken together, our study indicated that targeting NDUFC1 could open innovative perspectives for new multi-targeting approaches in the treatment of gastric cancer.
    Keywords:  NDUFC1; cell apoptosis; cell migration; cell proliferation; gastric cancer
    DOI:  https://doi.org/10.3389/fonc.2021.709044
  20. Lab Invest. 2021 Dec 28.
    Overexpression of carnitine palmitoyltransferase 1A promotes mitochondrial fusion and differentiation of glioblastoma stem cells.
    Min Luo, Yu-Qi Liu, Hua Zhang, Chun-Hua Luo, Qing Liu, Wen-Ying Wang, Zhi-Cheng He, Cong Chen, Xiao-Ning Zhang, Min Mao, Kai-Di Yang, Chao Wang, Xiao-Qing Chen, Wen-Juan Fu, Qin Niu, Xiu-Wu Bian, Yu Shi, Yi-Fang Ping.
      Glioma stem cells (GSCs) are self-renewing tumor cells with multi-lineage differentiation potential and the capacity of construct glioblastoma (GBM) heterogenicity. Mitochondrial morphology is associated with the metabolic plasticity of GBM cells. Previous studies have revealed distinct mitochondrial morphologies and metabolic phenotypes between GSCs and non-stem tumor cells (NSTCs), whereas the molecules regulating mitochondrial dynamics in GBM cells are largely unknown. Herein, we report that carnitine palmitoyltransferase 1A (CPT1A) is preferentially expressed in NSTCs, and governs mitochondrial dynamics and GSC differentiation. Expressions of CPT1A and GSC marker CD133 were mutually exclusive in human GBMs. Overexpression of CPT1A inhibited GSC self-renewal but promoted mitochondrial fusion. In contrast, disruption of CPT1A in NSTCs promoted mitochondrial fission and reprogrammed NSTCs toward GSC feature. Mechanistically, CPT1A overexpression increased the phosphorylation of dynamin-related protein 1 at Ser-637 to promote mitochondrial fusion. In vivo, CPT1A overexpression decreased the percentage of GSCs, impaired GSC-derived xenograft growth and prolonged tumor-bearing mice survival. Our work identified CPT1A as a critical regulator of mitochondrial dynamics and GSC differentiation, indicating that CPT1A could be developed as a molecular target for GBM cell-differentiation strategy.
    DOI:  https://doi.org/10.1038/s41374-021-00724-0
  21. Front Physiol. 2021 ;12 750627
    Redox-Sensitive VDAC: A Possible Function as an Environmental Stress Sensor Revealed by Bioinformatic Analysis.
    Andonis Karachitos, Wojciech Grabiński, Martyna Baranek, Hanna Kmita.
      Voltage-dependent anion-selective channel (VDAC) allows the exchange of small metabolites and inorganic ions across the mitochondrial outer membrane. It is involved in complex interactions that regulate mitochondrial and cellular functioning. Many organisms have several VDAC paralogs that play distinct but poorly understood roles in the life and death of cells. It is assumed that such a large diversity of VDAC-encoding genes might cause physiological plasticity to cope with abiotic and biotic stresses known to impact mitochondrial function. Moreover, cysteine residues in mammalian VDAC paralogs may contribute to the reduction-oxidation (redox) sensor function based on disulfide bond formation and elimination, resulting in redox-sensitive VDAC (rsVDAC). Therefore, we analyzed whether rsVDAC is possible when only one VDAC variant is present in mitochondria and whether all VDAC paralogs present in mitochondria could be rsVDAC, using representatives of currently available VDAC amino acid sequences. The obtained results indicate that rsVDAC can occur when only one VDAC variant is present in mitochondria; however, the possibility of all VDAC paralogs in mitochondria being rsVDAC is very low. Moreover, the presence of rsVDAC may correlate with habitat conditions as rsVDAC appears to be prevalent in parasites. Thus, the channel may mediate detection and adaptation to environmental conditions.
    Keywords:  VDAC; cysteine oxidation; environmental stress; parasite; redox sensor; spermatozoa
    DOI:  https://doi.org/10.3389/fphys.2021.750627
  22. J Biol Chem. 2021 Dec 28. pii: S0021-9258(21)01357-0. [Epub ahead of print] 101547
    A cell culture platform for quantifying metabolic substrate oxidation in bicarbonate-buffered medium.
    James R Krycer, Mary Lor, Rebecca L Fitzsimmons, James E Hudson.
      Complex diseases such as cancer and diabetes are underpinned by changes in metabolism, specifically by which and how nutrients are catabolized. Substrate utilization can be directly examined by measuring a metabolic endpoint rather than an intermediate (such as tricarboxylic cycle metabolite). For instance, oxidation of specific substrates can be measured in vitro by incubation of live cultures with substrates containing radiolabeled carbon and measuring radiolabeled carbon dioxide. To increase throughput, we previously developed a miniaturized platform to measure substrate oxidation of both adherent and suspension cells using multiwell plates rather than flasks. This enabled multiple conditions to be examined simultaneously, ideal for drug screens and mechanistic studies. However, like many metabolic assays, this was not compatible with bicarbonate-buffered media, which is susceptible to alkalinization upon exposure to gas containing little carbon dioxide such as air. While other buffers such as HEPES can overcome this problem, bicarbonate has additional biological roles as a metabolic substrate and in modulating hormone signaling. Here, we create a bicarbonate-buffered well-plate platform to measure substrate oxidation. This was achieved by introducing a sealed environment within each well that was equilibrated with carbon dioxide, enabling bicarbonate buffering. As proof of principle, we assessed metabolic flux in cultured adipocytes, demonstrating that bicarbonate-buffered medium increased lipogenesis, glucose oxidation, and sensitivity to insulin in comparison to HEPES-buffered medium. This convenient and high-throughput method facilitates the study and screening of metabolic activity under more physiological conditions to aid biomedical research.
    Keywords:  Gas trap; adipocyte; bicarbonate; carbon dioxide; cell metabolism; glucose; oxidation
    DOI:  https://doi.org/10.1016/j.jbc.2021.101547
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